Running exercise with and without calcium supplementation from tuna bone reduced bone impairment caused by low calcium intake in young adult rats.

Inadequate calcium intake during childhood and adolescence is detrimental to bone metabolism. Here, we postulated that calcium supplement prepared from tuna bone with tuna head oil should benefit for skeletal development than CaCO3. Forty female 4-week-old rats were divided into calcium-replete diet (0.55% w/w, S1, n = 8) and low-calcium groups (0.15% w/w for 2 weeks; L; n = 32). Then L were subdivided into 4 groups (8/group), i.e., remained on L, L + tuna bone (S2), S2 + tuna head oil + 25(OH)D3 and S2 + 25(OH)D3. Bone specimens were collected at week 9. We found that 2 weeks on low calcium diet led to low bone mineral density (BMD), reduced mineral content, and impaired mechanical properties in young growing rats. Intestinal fractional calcium absorption also increased, presumably resulting from higher plasma 1,25(OH)2D3 (1.712 ± 0.158 in L vs. 1.214 ± 0.105 nM in S1, P < 0.05). Four-week calcium supplementation from tuna bone further increased calcium absorption efficacy, which later returned to the basal level by week 9. Calcium supplementation successfully restored BMD, bone strength and microstructure. However, 25(OH)D3 + tuna head oil + tuna bone showed no additive effect. Voluntary running also effectively prevented bone defects. In conclusion, both tuna bone calcium supplementation and exercise are effective interventions for mitigating calcium-deficient bone loss.

Comparison of neuroprotective effects of dihydrotestosterone, 17ß-estradiol, and Pueraria mirifica herb extract on cognitive impairment in androgen deficient male rats.

This study investigated the neuroprotective effects of dihydrotestosterone (DHT), 17ß-estradiol (E2), and Pueraria mirifica herb extract (PME; an alternative source of natural estrogens) on the (i) learning and memory in androgen-deficient male rats, and on the hippocampus expression levels of (ii) mRNA of genes associated with synaptic transmission and structure, neurofibrillary tangles, and amyloid plaques, and (iii) total and phosphorylated tau proteins. The four-month-old male rats were sham-operated or orchidectomized (ODX). The ODX rats were divided into four groups, and orally treated for 2 months with either 1 mL/d of distilled water or 100 mg/kg/d of PME; or subcutaneously injected with 1 mg/kg/d of DHT or 80 µg/kg/d of E2. The impairment of spatial learning behavior and memory capacity in the ODX rats was prevented by DHT, E2, and PME. Recovery of the orchidectomy-induced deterioration of the synaptic plasticity in the hippocampus of rats was ranked as E2 ≥ PME > DHT. Both DHT and PME mitigated the increased Tau3 and Tau4 mRNA levels, and Tau-5 and P-Tau Ser396 protein levels more than E2 (DHT ≥ PME > E2). Only DHT tended to decrease App mRNA expression level. In conclusion, DHT showed a stronger efficacy for mitigation of the impaired spatial learning behavior and memory capacity in androgen-deficient male rats compared to E2 and PME, and their mechanisms of action are slightly different.

Comparative Pharmacokinetics of Puerarin Alone and in Pueraria mirifica Extract in Female Cynomolgus Monkeys.

Pueraria mirifica is an endemic Thai plant that has been used for rejuvenation and in the relief of various aging diseases. Puerarin is one of the major isoflavones found in this plant and shows several pharmacological activities in relation to the Thai traditional use of P. mirifica. Therefore, comparative pharmacokinetics of pure puerarin alone and that in a P. mirifica extract in cynomolgus monkeys were conducted in order to investigate the pharmacokinetic profiles of the 2 preparations. To this end, puerarin and P. mirifica extract, at an equivalent dose of 10 mg/kg of puerarin, were orally dosed to adult female monkeys for 7 consecutive days. A single intravenous injection of puerarin at a dose of 1 mg/kg was also peformed. Serial blood samples and excreta were collected from 0 - 24 h and 0 - 48 h after dosing. Determination of the puerarin levels and its metabolites in biological samples was conducted by liquid chromatography tandem mass spectrometry. Plasma levels of aspartate aminotransferase, alanine aminotransferase, and creatinine fluctuated in the normal range, with no abnormal physical signs in the animal. The absolute oral bioavailability of puerarin was approximately 1% in both preparations. Accumulation of puerarin was found after oral dosing for 7 consecutive days in both groups. Major metabolites of puerarin found in monkeys were hydroxylation and deglycosylation products. A negligible amount of unchanged puerarin was detected in urine and feces. Pharmacokinetic profiles obtained from this study could help to design the prescribed remedy of puerarin and P. mirifica extract phytopharmaceutical products for human use.

Absolute oral bioavailability and disposition kinetics of puerarin in female rats.

BACKGROUND: Pueraria candollei var. mirifica is a medicinal plant that is promoted as a "Champion Product" by the Government of Thailand. This plant has been reported to relieve postmenopausal symptoms, prevent and reverse bone loss, inhibit the growth of breast cancer, and alleviate cardiovascular diseases in preclinical and clinical studies. However, there is little information on the oral bioavailability and tissue distribution of puerarin with respect to its pharmacodynamic activities. Therefore, the aim of this study was to determine the pharmacokinetics of puerarin, including absorption, distribution, metabolism, and elimination, in rats. Moreover, this is the first study to examine the tissue distribution of puerarin in the hippocampus, femur, tibia, and mammary gland. METHODS: Adult female rats were administered puerarin at 1 mg/kg intravenously or 5 and 10 mg/kg orally. Blood, tissue, urine, and feces were collected and analyzed by liquid chromatography-tandem mass spectrometry. RESULTS: Puerarin reached a maximum concentration in the blood of 140-230 µg/L within 1 h of oral dosing, and had an absolute oral bioavailability of approximately 7%. Following intravenous administration, puerarin was widely distributed in several tissues, including the hippocampus, heart, lung, stomach, liver, mammary gland, kidney, spleen, femur, and tibia. Approximately 50% of the intravenous dose was excreted as glucuronide metabolites via the urinary route. CONCLUSIONS: The absolute oral bioavailability of puerarin was approximately 7% at doses of 5 and 10 mg/kg. Puerarin was widely distributed to several organs related to the diseases of aging, including the hippocampus, femur, tibia, and mammary gland. Glucuronides were the major metabolites of puerarin and were mainly excreted in the urine. These results are useful for the development of puerarin and Pueraria candollei var. mirifica as phytopharmaceutical products.

Pueraria mirifica alleviates cortical bone loss in naturally menopausal monkeys.

Since the in vitro and in vivo anti-osteoporotic effects of Pueraria mirifica (PM) in rodents have been verified, its activity in menopausal monkeys was evaluated as required before it can be applicable for human use. In this study, postmenopausal osteoporotic monkeys were divided into two groups (five per group), and fed daily with standard diet alone (PMP0 group) or diet mixed with 1000 mg/kg body weight (BW) of PM powder (PMP1000 group) for 16 months. Every 2 months, the bone mineral density (BMD), bone mineral content (BMC) and bone geometry parameters (cortical area and thickness and periosteal and endosteal circumference) at the distal radius and proximal tibia were determined using peripheral quantitative computed tomography together with plasma and urinary bone markers. Compared with the baseline (month 0) values, the cortical, but not trabecular, BMDs and BMCs and the cortical area and thickness at the metaphysis and diaphysis of the radius and tibia of the PMP0 group continuously decreased during the 16-month study period. In contrast, PMP1000 treatment ameliorated the bone loss mainly at the cortical diaphysis by decreasing bone turnover, as indicated by the lowered plasma bone-specific alkaline phosphatase and osteocalcin levels. Generally, changes in the cortical bone geometry were in the opposite direction to the cortical bone mass after PMP1000 treatment. This study indicated that postmenopausal monkeys continuously lose their cortical bone compartment, and they have a higher possibility for long bone fractures. Oral PMP treatment could improve both the bone quantity (BMC and BMD) and quality (bone geometry).

Neurotherapeutic Effects of Pueraria mirifica Extract in Early- and Late-Stage Cognitive Impaired Rats.

We determined the neurotherapeutic effects of Pueraria mirifica extract (PME) and pure puerarin (PU) in comparison with 17ß-estradiol (E2 ) in early- and late-stage cognitive impaired rats. Rats were ovariectomized (OVX), kept for 2 and 4 months to induce early- and late-stage cognitive impairment, respectively, and divided into four groups that were treated daily with (i) distilled water, (ii) 100 mg/kg of PME, (iii) 7 mg/kg of PU, and (iv) 80 µg/kg of E2 for 4 months. The estrogen deficiency symptoms of OVX rats were abrogated by treatment with E2 or PME, but not by treatment with PU. The mRNA level of genes associated with amyloid production (App and Bace1) and hyperphosphorylated Tau (Tau4) were upregulated together with the level of impaired cognition in the 2- and 4-month OVX rats. Treatment with E2 reduced the level of cognitive impairment more than that with PME and PU, and 2-month OVX rats were more responsive than 4-month OVX rats. All treatments down-regulated the Bace1 mRNA level in 2-month OVX rats, while PU and PME also decreased the App mRNA level in 2- and 4-month OVX rats, respectively. Only PU suppressed Tau4 expression in 2-month OVX rats. Thus, PME and PU elicit neurotherapeutic effects in different pathways, and earlier treatment is optimal. Copyright © 2016 John Wiley & Sons, Ltd.

Anti-osteoporotic effects of Pueraria candollei var. mirifica on bone mineral density and histomorphometry in estrogen-deficient rats.

Although it has been clearly shown that Pueraria mirifica and its phytoestrogens can mimic estrogen in preventing bone loss, as osteoporosis is an asymptomatic disease, the therapeutic effects of P. mirifica should be acknowledged. In this study, 6-month-old female rats were ovariectomized, kept for 4 weeks to induce bone loss, divided into five groups, and treated with P. mirifica at doses of 0, 5, 25, and 50 mg/kg BW/day (PM0, PM5, PM25, and PM50 groups, respectively) or 7 mg/kg BW/day of puerarin (PU group) for 12 weeks. Only the trabecular bone mineral densities (BMDs) of tibia metaphysis (at the 12th, 14th, and 16th week) and total and trabecular BMDs of L4 (at the 16th week) of the PM50 group were significantly higher than those of the PM0 group. However, the BMDs of tibia metaphysis and L4 at the 16th week of the study period were kept significantly lower than those of the 0 week, and the BMD was also significantly lower than that of the 4th week for tibia metaphysis. The trabecular bone area (BV/TV), trabecular number (Tb.N), and osteoblast surface (Ob.S/BS) were significantly higher, and trabecular space (Tb.Sp) was significantly lower in the PM50 group, as compared with those of the PM0 group. This study indicates that P. mirifica could be used as an anti-osteoporotic agent for postmenopausal women. Since P. mirifica could mainly retain bone mass at the levels before bone loss is initiated, the use of other anabolic agents in combination with P. mirifica is recommended for osteoporotic patients.

Pueraria mirifica extract and puerarin enhance proliferation and expression of alkaline phosphatase and type I collagen in primary baboon osteoblasts.

Phytoestrogen-rich Pueraria mirifica (PM) tuberous extract is a promising candidate for the development of anti-osteoporosis drugs for postmenopausal women, but its action has never been validated in humans or in non-human primates, which are more closely related to humans than rodents. In vitro study of non-human primate osteoblasts is thus fundamental to prepare for in vivo studies of phytoestrogen effects on primate bone. This study aimed to establish a culture system of baboon primary osteoblasts and to investigate the effects of PM extract and its phytoestrogens on these cells. Primary osteoblasts from adult baboon fibulae exhibited osteoblast characteristics in regard to proliferation, differentiation, mineralization, and estrogen receptor expression. They responded to 17ß-estradiol by increased proliferation rate and mRNA levels of alkaline phosphatase (ALP), type I collagen, and osteocalcin. After being exposed for 48 h to 100 µg/ml PM extract, 1000 nM genistein, or 1000 nM puerarin, primary baboon osteoblasts markedly increased the rate of proliferation and mRNA levels of ALP and type I collagen without changes in Runx2, osterix, or osteocalcin expression. PM extract, genistein, and puerarin also decreased the RANKL/OPG ratio, suggesting that they could decrease osteoclast-mediated bone resorption. However, neither PM extract nor its phytoestrogens altered calcium deposition in osteoblast culture. In conclusion, we have established baboon primary osteoblast culture, which is a new tool for bone research and drug discovery. Furthermore, the present results provide substantial support for the potential of PM extract and its phytoestrogens to be developed as therapeutic agents against bone fragility.

Improvements of vaginal atrophy without systemic side effects after topical application of Pueraria mirifica, a phytoestrogen-rich herb, in postmenopausal cynomolgus macaques.

The estrogenic efficacy of topical vaginal application of Pueraria mirifica extract (PM) on the restoration of vaginal atrophy, and the presence of any systemic side effects, were investigated in postmenopausal cynomolgus macaques. Twelve postmenopausal cynomolgus macaques, with complete cessation of menstruation for at least 5 years before start of this experiment, were divided into three groups. They received a topical vaginal application daily of 0.1 or 1% (w/w) PM cream or a conjugated equine estrogen (CEE) cream (a mixture of estrone, equilin, 17ß-dihydroequilin, 17α-estradiol and 17α-dihydroequilin at 0.625 mg total estrogen/g cream) for 28 days. Estrogenic efficacy was assessed weekly by vaginal cytology assay and vaginal pH measurement, whilst the plasma luteinizing hormone (LH) and sex skin coloration levels were determined at the end of each treatment period to evaluate the systemic side effects. PM significantly increased the proportion of superficial cells in a dose-dependent manner, with a similar efficacy between 1% (w/w) PM and CEE. Together with increased vaginal maturation, PM decreased the vaginal pH to acidic levels, as observed in the CEE group. PM induced no detected systemic side effects, whilst CEE decreased the plasma LH level and increased the reddish color of the sex skin during the posttreatment period. Topical vaginal treatment with PM stimulated the maturation of the vaginal epithelium without causing systemic side effects in postmenopausal monkeys. The implication is that PM could be a safer alternative to treat vaginal atrophy in postmenopausal women.

Upregulation of osteoblastic differentiation marker mRNA expression in osteoblast-like UMR106 cells by puerarin and phytoestrogens from Pueraria mirifica.

Phytoestrogens have attracted attention for their potential in the prevention of postmenopausal osteoporosis. Recently, phytoestrogen-rich herb Pueraria mirifica has been demonstrated to possess an osteogenic effect on bone in ovariectomized rats, but its underlying cellular mechanism was not known. Here, we investigated the effects of P. mirifica extract and its major isoflavone compound, puerarin, on cell viability, cell proliferation and the expression of differentiation markers in rat osteoblast-like UMR106 cells. After exposure to 17ß-estradiol (E2), genistein, P. mirifica extract and puerarin, proliferation but not viability of UMR106 cells was markedly decreased. Quantitative real-time PCR revealed that P. mirifica extract and puerarin significantly increased the mRNA expression of alkaline phosphatase (ALP) and osteoprotegerin, but not Runx2, osterix or osteocalcin. Puerarin also decreased the mRNA expression of receptor activator of nuclear factor-κB ligand, an osteoclastogenic factor, suggesting that it could induce bone gain by enhancing osteoblast differentiation and suppressing osteoclast function. Furthermore, after an exposure to high affinity estrogen receptor (ER) antagonist (ICI182780), the E2-, genistein-, P. mirifica extract- and puerarin-induced upregulation of ALP expressions were completely abolished. It could be concluded that P. mirifica extract and puerarin induced osteoblast differentiation rather than osteoblast proliferation in an ER-dependent manner. The present findings, therefore, corroborated the potential benefit of P. mirifica extract and puerarin in the prevention and treatment of postmenopausal osteoporosis.

Testing of the estrogenic activity and toxicity of Stephania venosa herb in ovariectomized rats.

Stephania venosa Spreng is a traditional herb which has been used for cancer treatment as well as an aphrodisiac. The scientific literature strongly supports its in vitro antiproliferative effects on cancer cell lines and has suggested developing this plant as a potential anticancer drug. However, the in vivo steroidogenic activity and toxicity of this plant have never been tested. We analyzed the levels of five key isoflavones in the plant extract by quantitative HPLC and then evaluated the in vivo estrogenic activity and toxicity in ovariectomized rats, in comparison with the phytoestrogen-rich plant, Pueraria mirifica. Twenty rats were first ovariectomized, and then seven days later divided into four groups and gavaged daily with 0, 10 and 100 mg/kg body weight/day of S. venosa, or 100 mg/kg body weight/day of P. mirifica for 28 days. A trace amount of puerarin, daidzin and daidzein with a subtle amount of genistein and genistin were isolated from the S. venosa tuber extract. S. venosa tuber powder, at both doses, did not exhibit any detectable estrogenic activity in ovariectomized rats, as assessed by the vaginal cytology and uterotropic assays, whilst P. mirifica induced a remarkable vaginal and uterine proliferation. S. venosa induced a toxicological effect on the hematological values and histopathological appearance of metabolic organs. Taken together, these results suggest that S. venosa has no discernable estrogenic activity but that it is toxic, at least to ovariectomized rats. Thus, the use of this plant for anticancer treatment needs to be reassessed or used with caution.

Medical applications of phytoestrogens from the Thai herb Pueraria mirifica.

Pueraria mirifica Airy Shaw et Suvatabandhu is a medicinal plant endemic to Thailand. It has been used in Thai folklore medicine for its rejuvenating qualities in aged women and men for nearly one hundred years. Indeed, it has been claimed that P. mirifica contains active phytoestrogens (plant substances with estrogen-like activity). Using high performance liquid chromatography, at least 17 phytoestrogens, mainly isoflavones, have been isolated. Thus, fairly considerable scientific researches, both in vitro in cell lines and in vivo in various species of animals including humans, have been conducted to date to address its estrogenic activity on the reproductive organs, bones, cardiovascular diseases and other climacteric related symptoms. The antioxidative capacity and antiproliferative effect on tumor cell lines have also been assessed. In general, P. mirifica could be applicable for preventing, or as a therapeutic for, the symptoms related to estrogen deficiency in menopausal women as well as in andropausal men. However, the optimal doses for each desirable effect and the balance to avoid undesired side effects need to be calculated before use.

Puerarin exhibits weak estrogenic activity in female rats.

A weak estrogenicity of puerarin on reproductive organs was addressed in female rats. In short-term treatment, immature ovariectomized rats were injected with 0.7 mg/kg BW/day of puerarin, for 14 days. Puerarin did not increase uterus weights, endometrium and myometrium areas, and the percent of cornified cells (%Co), but it increased the number of uterine glands. In long-term treatment, mature rats were injected with 7.0mg/kg BW/day of puerarin for 140 days. Puerarin did not increase uterus weights, endometrium and myometrium areas, and the number of uterine glands, but a significant increase in the %Co was observed from day 98 onwards.

Androgenic activity of the Thai traditional male potency herb, Butea superba Roxb., in female rats.

AIM OF THE STUDY: Butea superba Roxb. (Leguminosae) is a well-known Thai male potency herb with androgenic and anti-estrogenic activities. We evaluated whether oral administration of Butea superba has an androgenic or anti-estrogenic activity in female rats. MATERIALS AND METHODS: Normal and ovariectomized adult female rats were each subdivided into five groups, DW, BS-10, BS-50, BS-250 and TP, and gavaged with 0, 10, 50 and 250 mg/kg BW/day of the crude of Butea superba and subcutaneously injected with 6 mg/kg BW/day of testosterone propionate (TP), respectively, during the treatment period. RESULTS: In intact rats, only BS-250 increased the uterine thickness and the number of uterine glands, and could induce a prolonged diestrous phase. In ovariectomized rats, treatment with BS-50 as well as BS-250 increased the uterine thickness and the number of uterine glands. However, serum luteinizing hormone (LH) levels were also increased. TP reduced serum follicle stimulating hormone and LH levels with the appearance of anestrous cycle, and could significantly increase the relative uterine weight and thickness and the number of uterine glands in both intact and ovariectomized rats. CONCLUSIONS: Orally administered Butea superba tubers have an androgenic effect on the reproductive organs of intact and ovariectomized rats, and exhibit anti-estrogenic activity on LH secretion in ovariectomized rats.

Variance of estrogenic activity of the phytoestrogen-rich plant.

OBJECTIVE: To evaluate the influences of seasonal changes and plant cultivars on estrogenic activity of the phytoestrogen-rich plant, Pueraria mirifica. METHODS: Three cultivars of P. mirifica; PM-I, PM-II and PM-V, were grown in the same field trial for 3 years and random tubers collected during the summer, rainy season and winter seasons. Female Wistar rats were ovariectomized, kept for 14 days, randomly segregated into groups and treated with one of DW, 200microg/100g BW 17beta-estradiol (E2) or tuberous powder of PM-I, PM-II and PM-V at dosages of 100, or 1000mg/kg BW for the next 14 days. For the last 7 days of post-treatment period, rats received only DW. The vaginal cornification was recorded during the treatment and post-treatment period. The uterine tissues of the treated rats at the treatment and post-treatment periods were analyzed for uterine gland number and for the surface area of the myometrium, endometrium and lumen. In addition, ethanol tuberous extracts of PM-I, PM-II and PM-V was submitted to DPPH analysis. RESULTS: Vaginal cornification exhibited a dose-dependent response with plant samples collected during the winter and summer being more active than those collected in the rainy season. All plant samples-induced uterotrophic effects in the analysis at the treatment and post-treatment periods in a dose-dependent manner. The P. mirifica treated rats exhibited increasing uterine gland numbers and thickness of the endometrium and myometrium but a decreasing size of lumen, in comparison to the negative control. The results were more prominent in PM-I than other plants and also in plant samples collected during the winter and summer seasons than in the rainy season. DPPH assay of the ethanol tuberous extracts revealed variance in antioxidant activity. CONCLUSION: The results of uterotrophic and vaginal cornification assays reveal that P. mirifica exhibits a dose-dependent estrogenic activity under the influence of both seasonal changes and plant cultivars, which is confirmed by DPPH assay.

Oral administration of Kaempferia parviflora did not disturb male reproduction in rats.

To investigate the androgenic effect of Kaempferia parviflora (KP), a Thai herbal plant, adult male rats were randomized into control and KP-treatment groups. Rats were treated orally with water in the control group and with 1,000 mg/kg/day of KP in the treatment group for 45 days. Blood samples were collected on days 10, 20, 30 and 45 for measurement of the serum follicle stimulating hormone (FSH), luteinizing hormone (LH), testosterone, progesterone and corticosterone levels. The reproductive and non-reproductive organs were dissected on day 45 and weighed. Mating behavior was also observed on days 20 and 30. Body weight was measured throughout the study period. The results showed that KP induced an increase in body weight compared with the controls. There were no significant differences in the weights of either reproductive (testis, seminal vesicle plus coagulating gland, levator ani muscle plus bulbocarvernosus muscle and glans penis, except the prostate gland) or non-reproductive organs (kidney, adrenal gland and gastracnemius muscle). There were no significant differences in serum levels of either FSH or LH between the two groups. The serum testosterone and progesterone levels were insignificantly lower in the KP group during the first 30 days. The serum corticosterone levels in the KP group were lower than those in the controls throughout the study period and were significantly low on days 20 and 30. There were no significant changes in mating behavior in the rats treated with KP. Although KP affected the body weight and serum corticosterone level, it did not affect mating behavior, reproductive and non-reproductive organ weights or hormones related to the reproductive system in the adult male rats. Therefore, we conclude that the testosterone-like effect of KP did not disturb the hypothalamic-pituitary-testicular axis or male reproduction.

Androgen disruption and toxicity tests of Butea superba Roxb., a traditional herb used for treatment of erectile dysfunction, in male rats.

OBJECTIVE: To evaluate the effects of the tuberous powder of Butea superba Roxb. (Leguminosae) on blood testosterone and luteinizing hormone (LH), and toxicity in male rats. METHODS: Adult male Wistar rats were orally treated with 0, 10, 100, 150 or 200 mg/kg BW/day of B. superba powder suspension in 0.7 ml distilled water for 90 consecutive days. Blood samples were collected every 30 days and submitted to testosterone and LH analysis. On the 90th day of treatment, blood and the main organs were collected for haematological and histopathological analysis, respectively. RESULTS: The adverse effects found included an increase in spleen relative weight, and increased serum level of alkaline phosphatase (ALP) and aspartate aminotransferase (AST) in rats treated with 150 mg/kg BW/day B. superba powder. At 200 mg/kg BW/day treatment, rats showed significant decreased and increased blood levels of neutrophil and eosinophil, respectively, and a decrease in serum creatinine levels. Serum hormonal analysis revealed a dose-dependent decrease in testosterone, but not LH, in rats treated with 150 and 200 mg/kg BW/day B. superba powder. CONCLUSION: Subchronic treatment of B. superba tuberous powder suspension at high doses in male rats exhibited adverse effects to blood chemistry, haematology, and blood testosterone level. The results of the study should initiate awareness of the possible adverse risk of over-dose consumption of B. superba products for treatment of erectile dysfunction (ED) in mature males.

Preventive effects of Pueraria mirifica on bone loss in ovariectomized rats.

OBJECTIVE: Effects of Pueraria mirifica on bone loss in fully mature ovariectomized rats are examined. METHODS: Two series of experiments were performed. In the first series, rats were kept with their ovaries intact and divided into two groups; initial control (IC) and sham control (SH). The IC rats were sacrificed on day 1 and their data were kept as baseline control. The SH rats were subjected to sham operation on day 0 and gavaged daily with distilled water for 90 days. In the second series, rats were subjected to ovariectomy, divided into five groups and gavaged daily with 0.1mg/kg B.W./day of 17-alpha-ethinylestradiol (EE), 0, 10, 100 and 1000mg/kg B.W./day of P. mirifica (P0, P10, P100 and P1000, respectively) for 90 days. Changes of bone mineral density and bone mineral content were measured using peripheral Quantitative Computerized Tomography. RESULTS: Bone loss was significantly induced by ovariectomy and it was dose-dependently prevented by P. mirifica treatment for 90 days. The preventive effects of P. mirifica on bone loss depended on bone types (axial or long bone), bone sites (metaphysis or diaphysis), and bone compartments (trabecular and cortical). At P100 and P1000, bone loss was completely prevented both in trabecular bone mineral density and content. The effects of P. mirifica were, as expected, comparable to that in the EE group. CONCLUSION: These results suggest that P. mirifica may be applicable to treat the osteoporosis in menopausal women; however, an undesirable side effect on stimulating reproductive organs should be concerned.

Assessment of urinary gonadotropin and steroid hormone profiles of female cynomolgus monkeys after treatment with Pueraria mirifica.

This study investigated the changes in the urinary hormone levels of female monkeys (Macaca fascicularis) after single-dose and long-term treatments with Pueraria mirifica (PM). The monkeys were separated into 3 groups (n=3) and orally treated with 10, 100, or 1,000 mg of PM in each group. Two series of experiments were performed. In the first series of experiments, the monkeys were orally treated with a single dose of PM. The experimental schedule was divided into a one menstrual cycle pretreatment period and a two menstrual cycle post-treatment period. In the second series of experiments, the monkeys were orally treated daily with PM for 90 days. The experiment schedule was divided into a one menstrual cycle pretreatment period, a three menstrual cycle treatment period, and a two menstrual cycle post-treatment period. Urinary samples were collected daily and assayed for the FSH, LH, estradiol, and progesterone levels. The results showed that there were no changes in the FSH, LH, estradiol, and progesterone levels after treatment with a single dose of 10, 100, or 1,000 mg of PM or after daily treatment with 10 mg of PM for 90 days compared with the levels observed during the pretreatment period. Daily treatment with 100 mg and 1,000 mg of PM for 90 days only produced a clear reduction in the urinary FSH levels. This suggests that changes of urinary FSH levels can be considered an indicator for study of estrogenic effects on hormonal levels in female monkeys.